RNA-seqに基づくmiRNAシグネチャーは小児B細胞性急性リンパ性白血病患者の独立した再発予測因子となる

京都大学新制・論文博士博士(医学)乙第13609号論医博第2319号京都大学大学院医学研究科医学専攻(主査)教授 村川 泰裕, 教授 竹内 理, 教授 永井 純正学位規則第4条第2項該当Doctor of Medical ScienceKyoto UniversityDFA

Age, gender, insulin and blood glucose control status alter the risk of ischemic heart disease and stroke among elderly diabetic patients

<p>Abstract</p> <p>Background</p> <p>We analyzed the effects of insulin therapy, age and gender on the risk of ischemic heart disease (IHD) and cerebrovascular accident (CVA) according to glycemic control.</p> <p>Methods and Results</p> <p>We performed a prospective cohort study (Japan Cholesterol and Diabetes Mellitus Study) of type 2 diabetes patients (n = 4014) for 2 years. The primary endpoint was the onset of fatal/non-fatal IHD and/or CVA, which occurred at rates of 7.9 and 7.2 per 1000 person-years, respectively. We divided diabetic patients into four groups based on age (≤ 70 and > 70) and hemoglobin A1C levels (≤ 7.0 and > 7.0%). Multiple regression analysis revealed that IHD was associated with high systolic blood pressure and low HDL-C in patients under 70 years of age with fair glycemic control and was associated with low diastolic blood pressure in the older/fair group. Interestingly, insulin use was associated with IHD in the older/poor group (OR = 2.27, 95% CI = 1.11-5.89; p = 0.026) and was associated with CVA in the older/fair group (OR = 2.09, 95% CI = 1.06-4.25; p = 0.028). CVA was associated with lower HDL-C and longer duration of diabetes in younger/poor glycemic control group. Results by stepwise analysis were similar. Next, patients were divided into four groups based on gender and diabetic control(hemoglobinA1C < or > 7.0%). Multiple regression analysis revealed that IHD was associated with high systolic blood pressure in male/fair glycemic control group, age in male/poor control group, and short duration of diabetic history in females in both glycemic control groups. Interestingly, insulin use was associated with IHD in the male/poor group(OR = 4.11, 95% CI = 1.22-8.12; p = 0.018) and with CVA in the female/poor group(OR = 3.26, 95% CI = 1.12-6.24; p = 0.02). CVA was associated with short duration of diabetes in both female groups.</p> <p>Conclusions</p> <p>IHD and CVA risks are affected by specific factors in diabetics, such as treatment, gender and age. Specifically, insulin use has a potential role in preventing IHD but may also be a risk factor for CVA among the diabetic elderly, thus revealing a need to develop improved treatment strategies for diabetes in elderly patients. The Japan Cholesterol and Diabetes Mellitus Study was formulated to evaluate them(Umin Clinical Trials Registry, clinical trial reg. no. UMIN00000516; <url>http://www.umin.ac.jp/ctr/index.htm</url>).</p

RUNX inhibitor suppresses graft‐versus‐host disease through targeting RUNX‐NFATC2 axis

Patients with refractory graft-versus-host disease (GVHD) have a dismal prognosis. Therefore, novel therapeutic targets are still needed to be identified. Runt-related transcriptional factor (RUNX) family transcription factors are essential transcription factors that mediate the essential roles in effector T cells. However, whether RUNX targeting can suppress, and GVHD is yet unknown. Here, we showed that RUNX family members have a redundant role in directly transactivating NFATC2 expression in T cells. We also found that our novel RUNX inhibitor, Chb-M’, which is the inhibitor that switches off the entire RUNX family by alkylating agent–conjugated pyrrole-imidazole (PI) polyamides, inhibited T-cell receptor mediated T cell proliferation and allogenic T cell response. These were designed to specifically bind to consensus RUNX-binding sequences (TGTGGT). Chb-M’ also suppressed the expression of NFATC2 and pro-inflammatory cytokine genes in vitro. Using xenogeneic GVHD model, mice injected by Chb-M’ showed almost no sign of GVHD. Especially, the CD4 T cell was decreased and GVHD-associated cytokines including tissue necrosis factor-α and granulocyte-macrophage colony-stimulating factor were reduced in the peripheral blood of Chb-M’ injected mice. Taken together, our data demonstrates that RUNX family transcriptionally upregulates NFATC2 in T cells, and RUNX-NFATC2 axis can be a novel therapeutic target against GVHD

PAX5 alterations in an infant case of KMT2A-rearranged leukemia with lineage switch

Lineage switch is a rare event at leukemic relapse. While mostly known to occur in KMT2A-rearranged infant leukemia, the underlying mechanism is yet to be depicted. This case report describes a female infant who achieved remission of KMT2A-MLLT3-rearranged acute monocytic leukemia, but 6 months thereafter, relapsed as KMT2A-MLLT3-rearranged acute lymphocytic leukemia. Whole exome sequencing of the bone marrow obtained pre-post lineage switch revealed two somatic mutations of PAX5 in the relapse sample. These two PAX5 alterations were suggested to be loss of function, thus to have played the driver role in the lineage switch from acute monocytic leukemia to acute lymphocytic leukemia

The first Japanese biobank of patient‐derived pediatric acute lymphoblastic leukemia xenograft models

A lack of practical resources in Japan has limited preclinical discovery and testing of therapies for pediatric relapsed and refractory acute lymphoblastic leukemia (ALL), which has poor outcomes. Here, we established 57 patient-derived xenografts (PDXs) in NOD.Cg-Prkdcscidll2rgtm1Sug/ShiJic (NOG) mice and created a biobank by preserving PDX cells including three extramedullary relapsed ALL PDXs. We demonstrated that our PDX mice and PDX cells mimicked the biological features of relapsed ALL and that PDX models reproduced treatment-mediated clonal selection. Our PDX biobank is a useful scientific resource for capturing drug sensitivity features of pediatric patients with ALL, providing an essential tool for the development of targeted therapies

A RUNX-targeted gene switch-off approach modulates the BIRC5/PIF1-p21 pathway and reduces glioblastoma growth in mice

Glioblastoma is the most common adult brain tumour, representing a high degree of malignancy. Transcription factors such as RUNX1 are believed to be involved in the malignancy of glioblastoma. RUNX1 functions as an oncogene or tumour suppressor gene with diverse target genes. Details of the effects of RUNX1 on the acquisition of malignancy in glioblastoma remain unclear. Here, we show that RUNX1 downregulates p21 by enhancing expressions of BIRC5 and PIF1, conferring anti-apoptotic properties on glioblastoma. A gene switch-off therapy using alkylating agent-conjugated pyrrole-imidazole polyamides, designed to fit the RUNX1 DNA groove, decreased expression levels of BIRC5 and PIF1 and induced apoptosis and cell cycle arrest via p21. The RUNX1-BIRC5/PIF1-p21 pathway appears to reflect refractory characteristics of glioblastoma and thus holds promise as a therapeutic target. RUNX gene switch-off therapy may represent a novel treatment for glioblastoma

Inhibition of tumor-stromal interaction through HGF/Met signaling by valproic acid

Hepatocyte growth factor (HGF), which is produced by surrounding stromal cells, including fibroblasts and endothelial cells, has been shown to be a significant factor responsible for cancer cell invasion mediated by tumor-stromal interactions. We found in this study that the anti-tumor agent valproic acid (VPA), a histone deacetylase (HDAC) inhibitor, strongly inhibited tumor-stromal interaction. VPA inhibited HGF production in fibroblasts induced by epidermal growth factor (EGF), platelet-derived growth factor, basic fibroblast growth factor, phorbol 12-myristate 13-acetate (PMA) and prostaglandin E-2 without any appreciable cytotoxic effect. Other HDAC inhibitors, including butyric acid and trichostatin A (TSA), showed similar inhibitory effects on HGF production stimulated by various inducers. Up-regulations of HGF gene expression induced by PMA and EGF were also suppressed by VPA and TSA. Furthermore, VPA significantly inhibited HGF-induced invasion of HepG2 hepatocellular carcinoma cells. VPA, however, did not affect the increases in phosphorylation of MAPK and Akt in HGF-treated HepG2 cells. These results demonstrated that VPA inhibited two critical processes of tumor-stromal interaction, induction of fibroblastic HGF production and HGF-induced invasion of HepG2 cells, and suggest that those activities serve for other anti-tumor mechanisms of VPA besides causing proliferation arrest, differentiation, and/or apoptosis of tumor cells

RUNX1 transactivates BCR-ABL1 expression in Philadelphia chromosome positive acute lymphoblastic leukemia

The emergence of tyrosine kinase inhibitors as part of a front-line treatment has greatly improved the clinical outcome of the patients with Ph⁺ acute lymphoblastic leukemia (ALL). However, a portion of them still become refractory to the therapy mainly through acquiring mutations in the BCR-ABL1 gene, necessitating a novel strategy to treat tyrosine kinase inhibitor (TKI)-resistant Ph⁺ ALL cases. In this report, we show evidence that RUNX1 transcription factor stringently controls the expression of BCR-ABL1, which can strategically be targeted by our novel RUNX inhibitor, Chb-M'. Through a series of in vitro experiments, we identified that RUNX1 binds to the promoter of BCR and directly transactivates BCR-ABL1 expression in Ph⁺ ALL cell lines. These cells showed significantly reduced expression of BCR-ABL1 with suppressed proliferation upon RUNX1 knockdown. Moreover, treatment with Chb-M' consistently downregulated the expression of BCR-ABL1 in these cells and this drug was highly effective even in an imatinib-resistant Ph⁺ ALL cell line. In good agreement with these findings, forced expression of BCR-ABL1 in these cells conferred relative resistance to Chb-M'. In addition, in vivo experiments with the Ph⁺ ALL patient-derived xenograft cells showed similar results. In summary, targeting RUNX1 therapeutically in Ph⁺ ALL cells may lead to overcoming TKI resistance through the transcriptional regulation of BCR-ABL1. Chb-M' could be a novel drug for patients with TKI-resistant refractory Ph⁺ ALL

Alteration of the immune environment in bone marrow from children with recurrent B cell precursor acute lymphoblastic leukemia

Due to the considerable success of cancer immunotherapy for leukemia, the tumor immune environment has become a focus of intense research; however, there are few reports on the dynamics of the tumor immune environment in leukemia. Here, we analyzed the tumor immune environment in pediatric B cell precursor acute lymphoblastic leukemia by analyzing serial bone marrow samples from nine patients with primary and recurrent disease by mass cytometry using 39 immunophenotype markers, and transcriptome analysis. High-dimensional single-cell mass cytometry analysis elucidated a dynamic shift of T cells from naïve to effector subsets, and clarified that, during relapse, the tumor immune environment comprised a T helper 1-polarized immune profile, together with an increased number of effector regulatory T cells. These results were confirmed in a validation cohort using conventional flow cytometry. Furthermore, RNA transcriptome analysis identified the upregulation of immune-related pathways in B cell precursor acute lymphoblastic leukemia cells during relapse, suggesting interaction with the surrounding environment. In conclusion, a tumor immune environment characterized by a T helper 1-polarized immune profile, with an increased number of effector regulatory T cells, could contribute to the pathophysiology of recurrent B cell precursor acute lymphoblastic leukemia. This information could contribute to the development of effective immunotherapeutic approaches against B cell precursor acute lymphoblastic leukemia relapse